Project Summary/Abstract Background: Across the globe, non-typhoidal serovars of the intracellular bacterial pathogen Salmonella enterica caused 153 million cases of gastroenteritis and 57,000 deaths in 2010. S. enterica enterica ser. Typhimurium, the most prevalent non-typhoidal Salmonella (NTS) serovar, creates a durable niche inside host cell vacuoles by injecting effector proteins that subvert host signal transduction pathways. Recently, S. Typhimurium has been shown to promote its intracellular survival and replication by activating the host transcription factor STAT3. While STAT3 is known to promote cell differentiation, survival, and proliferation in response to cytokines and growth factors, it is unknown how Salmonella drives STAT3 phosphorylation or how STAT3 renders host cells more permissive to S. Typhimurium. Goal/Preliminary Data: I aim to understand the mechanism and function of STAT3 phosphorylation during Salmonella infection. Recently, our lab discovered that an S. Typhimurium effector, SarA, physically associates with STAT3 and induces STAT3 phosphorylation. However, SarA lacks identifiable structural homology to any known kinase domain, so I hypothesize that SarA initiates a signaling pathway to phosphorylate STAT3. Further, S. Typhimurium ?sarA exhibits reduced intracellular replication in human cells and infected mice. This SarA-dependent reduction is recapitulated by knockdown or inhibition of host STAT3 during wild-type infection, indicating that SarA promotes virulence by activating STAT3. Hence, I hypothesize that STAT3- targets promote bacterial replication in the Salmonella-containing vacuole (SCV). Approach: I will determine how SarA-interacting partners, identified by proteomics, interact with phosphorylated SarA motifs to drive STAT3 tyrosine phosphorylation. In parallel, I will verify potential SarA to STAT3 signaling components or regulators that were identified through common human genetic variants associated with the level of STAT3-target activation. Finally, I will determine how activated STAT3 promotes S. Typhimurium intracellular replication, by ascertaining how SarA-dependent transcriptional changes, identified by transcriptomics analysis, promote intracellular bacterial replication. Significance: Uncovering how S. Typhimurium activates STAT3 to manipulate host transcription and improve its intracellular niche will enhance our understanding of bacteria-host interaction and the function of the STAT3 pathway, a crucial player in autoimmunity and oncogenesis.